The primary long-term objective of the present proposal is to develop a rapid new tandem mass spectrometric method for peptide and protein sequence analysis with sub-picomole sensitivity and improved scope of application compared with current MS methods. The successful accomplishment of this goal is likely to yield enormous savings in time, effort, and money in many different areas of biomedical research (e.g. elucidation of posttranslational modifications of proteins and the rapid generation of partial protein sequences for DNA probes) as well as to render feasible powerful new approaches to biomedical problems. Specifically it is proposed to construct and develop a novel tandem time-of-flight mass spectrometer that uses matrix-assisted laser desorption for the efficient production of intact gas phase peptide ions and pulsed energetic electron bombardment to produce a fragmentation spectrum from a chosen peptide ion species of interest. Of central importance to the current proposal is the development of an effective means for producing informative fragmentation of peptides by electron bombardment. Because few studies have been undertaken in this area, it is proposed to undertake a detailed investigation of the systematics of energetic electron-induced dissociation of protonated peptide ions. The performance of the new method will be carefully assessed by applying it to a number of current biological problems.
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